The present invention relates to a newly identified protein, 22406, a human pyridoxal-phosphate dependent enzyme family member. In particular, the invention relates to the 22406 polypeptide and encoding nucleic acid molecules, methods of detecting the 22406 polypeptide and encoding nucleic acid molecules, and methods of diagnosing and treating 22406-related disorders. Also provided are vectors, host cells, and recombinant methods for making and using the novel molecules.
The pyridoxal-phosphate dependent family of enzymes require the co-factor, pyridoxal-5xe2x80x2-phosphate (pyridoxal-phosphate), for catalytic activity. Pyridoxal-phosphate dependent enzymes (B6 enzymes) catalyze manifold reactions in the metabolism of amino acids. L- and D-serine dehydratase, threonine dehydratase, and serine racemase are a few of the members of this family of enzymes. In all of the members of the family, the pyridoxal-phosphate group is attached to a lysine residue. The sequence around this residue is sufficiently conserved to allow the derivation of a pattern specific to pyridoxal-phosphate dependent enzymes.
The pyridoxal-phosphate dependent family member, serine racemase, has been shown to catalyze the direct racemization of L-serine to D-serine with a requirement for pyridoxal 5xe2x80x2-phosphate (Wolosker et al. (1999) PNAS 96:721-725). The properties of this enzyme resemble those of bacterial racemases, suggesting that the biosynthetic pathway for D-amino acids is conserved from bacteria to mammalian brain.
It has been demonstrated that D-serine is the endogenous ligand for the glycine site of the glutamate N-methyl-D-aspartate (NMDA) receptor (Mothet et al. (2000) PNAS 97:4926-4931). The amino acid D-serine is synthesized and stored in glia rather than in neurons. Released glutamate acts on receptors on the protoplasmic astrocytes closely opposed to the synapse to release D-serine, which co-activates post-synaptic NMDA receptors together with glutamate. As D-serine is formed by serine racemase, inhibitors of this enzyme can be expected to reduce NMDA neurotransmission.
D-serine has been shown to modify behavioral changes associated with learning, memory, convulsion, anxiety, psychotomimetic induced abnormal behavior, cerebellar ataxia, and neurodengeneration. Inhibitors of serine racemase can be expected to quell anxiety and epilepsy and to prevent damage from stroke and certain neurodegenerative conditions including Alzheimer""s disease. On the other hand, stimulating serine racemase might improve schizophrenia symptoms, which are partly caused by depressed NMDA receptor function.
Accordingly, members of the pyridoxal-phosphate dependent enzyme class are a major target for drug action and development. Therefore, it is valuable to the field of pharmaceutical development to identify and characterize previously unknown serine racemases. The present invention advances the state of the art by providing a previously unidentified human pyridoxal-phosphate dependent serine racemace.
The present invention is based, in part, on the discovery of a novel human pyridoxal-phosphate dependent enzyme family member, referred to herein as xe2x80x9c22406xe2x80x9d. The nucleotide sequence of a cDNA encoding 22406 is shown in SEQ ID NO:1, and the amino acid sequence of a 22466 polypeptide is shown in SEQ ID NO:2. In addition, the nucleotide sequence of the coding region is depicted in SEQ ID NO:3.
Accordingly, in one aspect the invention features a nucleic acid molecule which encodes a 22406 protein or polypeptide, e.g., a biologically active portion of the 22406 protein. In a preferred embodiment, the isolated nucleic acid molecule encodes a polypeptide having the amino acid sequence of SEQ ID NO:2; In other embodiments, the invention provides an isolated 22406 nucleic acid molecule having the nucleotide sequence shown in SEQ ID NO:1 or SEQ ID NO:3. In still other embodiments, the invention provides nucleic acid molecules that are substantially identical (e.g., naturally occurring allelic variants) to the nucleotide sequence shown in SEQ ID NO:1 or SEQ ID NO:3. In other embodiments, the invention provides a nucleic acid molecule which hybridizes under stringent hybridization conditions to a nucleic acid molecule comprising the nucleotide sequence of SEQ ID NO:1 or SEQ ID NO:3, wherein the nucleic acid encodes a full length 22406 protein or an active fragment thereof.
In a related aspect, the invention further provides nucleic acid constructs which include a 22406 nucleic acid molecule described herein. In certain embodiments, the nucleic acid molecules of the invention are operatively linked to native or heterologous regulatory sequences. Also included, are vectors and host cells containing the 22406 nucleic acid molecules of the invention e.g., vectors and host cells suitable for producing 22406 nucleic acid molecules and polypeptides.
In another related aspect, the invention provides nucleic acid fragments suitable as primers or hybridization probes for the detection of 22406-encoding nucleic acids.
In still another related aspect, isolated nucleic acid molecules that are antisense to a 22406 encoding nucleic acid molecule are provided.
In another aspect, the invention features 22406 polypeptides, and biologically active or antigenic fragments thereof that are useful, e.g., as reagents or targets in assays applicable to treatment and diagnosis of 22406-mediated or -related disorders. In another embodiment, the invention provides 22406 polypeptides having a 22406 activity. Preferred polypeptides are 22406 proteins including at least one pyridoxal-phosphate dependent enzyme family member domain, and, preferably, having a 22406 activity, e.g., a 22406 activity as described herein.
In other embodiments, the invention provides 22406 polypeptides, e.g., a 22406 polypeptide having the amino acid sequence shown in SEQ ID NO:2; an amino acid sequence that is substantially identical to the amino acid sequence shown in SEQ ID NO:2; or an amino acid sequence encoded by a nucleic acid molecule having a nucleotide sequence which hybridizes under stringent hybridization conditions to a nucleic acid molecule comprising the nucleotide sequence of SEQ ID NO:1 or SEQ ID NO:3, wherein the nucleic acid encodes a full length 22406 protein or an active fragment thereof.
In a related aspect, the invention further provides nucleic acid constructs which include a 22406 nucleic acid molecule described herein.
In a related aspect, the invention provides 22406 polypeptides or fragments operatively linked to non-22406 polypeptides to form fusion proteins.
In another aspect, the invention features antibodies and antigen-binding fragments thereof, that react with, or more preferably specifically bind 22406 polypeptides.
In another aspect, the invention provides methods of screening for compounds that modulate the expression or activity of the 22406 polypeptides or nucleic acids.
In still another aspect, the invention provides a process for modulating 22406 polypeptide or nucleic acid expression or activity, e.g. using the screened compounds. In certain embodiments, the methods involve treatment of conditions related to aberrant activity or expression of the 22406 polypeptides or nucleic acids, such as conditions involving neurological disorders.
The invention also provides assays for determining the activity of or the presence or absence of 22406 polypeptides or nucleic acid molecules in a biological sample, including for disease diagnosis.
In further aspect the invention provides assays for determining the presence or absence of a genetic alteration in a 22406 polypeptide or nucleic acid molecule, including for disease diagnosis.